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1.
Cell Mol Life Sci ; 81(1): 11, 2023 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-38117357

RESUMO

Metabolic bone disorders and associated fragility fractures are major causes of disability and mortality worldwide and place an important financial burden on the global health systems. These disorders result from an unbalance between bone anabolic and resorptive processes and are characterized by different pathophysiological mechanisms. Drugs are available to treat bone metabolic pathologies, but they are either poorly effective or associated with undesired side effects that limit their use. The molecular mechanism underlying the most common metabolic bone disorders, and the availability, efficacy, and limitations of therapeutic options currently available are discussed here. A source for the unmet need of novel drugs to treat metabolic bone disorders is marine organisms, which produce natural osteoactive compounds of high pharmaceutical potential. In this review, we have inventoried the marine osteoactive compounds (MOCs) currently identified and spotted the groups of marine organisms with potential for MOC production. Finally, we briefly examine the availability of in vivo screening and validation tools for the study of MOCs.


Assuntos
Produtos Biológicos , Doenças Ósseas Metabólicas , Humanos , Produtos Biológicos/farmacologia
2.
Cell Mol Life Sci ; 80(10): 310, 2023 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-37777592

RESUMO

Skeletal disorders are problematic aspects for the aquaculture industry as skeletal deformities, which affect most species of farmed fish, increase production costs and affect fish welfare. Following recent findings that show the presence of osteoactive compounds in marine organisms, we evaluated the osteogenic and mineralogenic potential of commercially available microalgae strains Skeletonema costatum and Tetraselmis striata CTP4 in several fish systems. Ethanolic extracts increased extracellular matrix mineralization in gilthead seabream (Sparus aurata) bone-derived cell cultures and promoted osteoblastic differentiation in zebrafish (Danio rerio) larvae. Long-term dietary exposure to both extracts increased bone mineralization in zebrafish and upregulated the expression of genes involved in bone formation (sp7, col1a1a, oc1, and oc2), bone remodeling (acp5a), and antioxidant defenses (cat, sod1). Extracts also improved the skeletal status of zebrafish juveniles by reducing the incidence of skeletal anomalies. Our results indicate that both strains of microalgae contain osteogenic and mineralogenic compounds, and that ethanolic extracts have the potential for an application in the aquaculture sector as dietary supplements to support fish bone health. Future studies should also identify osteoactive compounds and establish whether they can be used in human health to broaden the therapeutic options for bone erosive disorders such as osteoporosis.


Assuntos
Microalgas , Dourada , Animais , Humanos , Osteogênese , Peixe-Zebra , Suplementos Nutricionais , Dourada/genética , Dourada/metabolismo
3.
Int J Mol Sci ; 24(4)2023 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-36834795

RESUMO

Ectopic calcification refers to the pathological accumulation of calcium ions in soft tissues and is often the result of a dysregulated action or disrupted function of proteins involved in extracellular matrix mineralization. While the mouse has traditionally been the go-to model organism for the study of pathologies associated with abnormal calcium deposition, many mouse mutants often have exacerbated phenotypes and die prematurely, limiting the understanding of the disease and the development of effective therapies. Since the mechanisms underlying ectopic calcification share some analogy with those of bone formation, the zebrafish (Danio rerio)-a well-established model for studying osteogenesis and mineralogenesis-has recently gained momentum as a model to study ectopic calcification disorders. In this review, we outline the mechanisms of ectopic mineralization in zebrafish, provide insights into zebrafish mutants that share phenotypic similarities with human pathological mineralization disorders, list the compounds capable of rescuing mutant phenotypes, and describe current methods to induce and characterize ectopic calcification in zebrafish.


Assuntos
Calcinose , Cálcio , Humanos , Camundongos , Animais , Cálcio/metabolismo , Peixe-Zebra/genética , Calcinose/metabolismo , Osteogênese , Matriz Extracelular/metabolismo , Cálcio da Dieta/metabolismo , Calcificação Fisiológica
4.
Proc Natl Acad Sci U S A ; 119(48): e2209231119, 2022 11 29.
Artigo em Inglês | MEDLINE | ID: mdl-36417434

RESUMO

The shaping of bone structures relies on various cell types and signaling pathways. Here, we use the zebrafish bifurcating fin rays during regeneration to investigate bone patterning. We found that the regenerating fin rays form via two mineralization fronts that undergo an osteoblast-dependent fusion/stitching until the branchpoint, and that bifurcation is not simply the splitting of one unit into two. We identified tartrate-resistant acid phosphatase-positive osteolytic tubular structures at the branchpoints, hereafter named osteolytic tubules (OLTs). Chemical inhibition of their bone-resorbing activity strongly impairs ray bifurcation, indicating that OLTs counteract the stitching process. Furthermore, by testing different osteoactive compounds, we show that the position of the branchpoint depends on the balance between bone mineralization and resorption activities. Overall, these findings provide a unique perspective on fin ray formation and bifurcation, and reveal a key role for OLTs in defining the proximo-distal position of the branchpoint.


Assuntos
Proteínas de Peixe-Zebra , Peixe-Zebra , Animais , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Osteoblastos/metabolismo , Transdução de Sinais , Osso e Ossos/metabolismo
5.
Pharmaceuticals (Basel) ; 15(8)2022 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-36015130

RESUMO

Bone disorders affect millions of people worldwide and treatments currently available often produce undesirable secondary effects or have limited efficacy. It is therefore of the utmost interest for patients to develop more efficient drugs with reduced off-target activities. In the long process of drug development, screening and preclinical validation have recently gained momentum with the increased use of zebrafish as a model organism to study pathological processes related to human bone disorders, and the development of zebrafish high-throughput screening assays to identify bone anabolic compounds. In this review, we provided a comprehensive overview of the literature on zebrafish bone-related assays and evaluated their performance towards an integration into screening pipelines for the discovery of mineralogenic/osteogenic compounds. Tools available to standardize fish housing and feeding procedures, synchronize embryo production, and automatize specimen sorting and image acquisition/analysis toward faster and more accurate screening outputs were also presented.

6.
Toxins (Basel) ; 14(7)2022 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-35878205

RESUMO

The dinoflagellate Amyloodinium ocellatum is the etiological agent of a parasitic disease named amyloodiniosis. Mortalities of diseased fish are usually attributed to anoxia, osmoregulatory impairment, or opportunistic bacterial infections. Nevertheless, the phylogenetic proximity of A. ocellatum to a group of toxin-producing dinoflagellates from Pfiesteria, Parvodinium and Paulsenella genera suggests that it may produce toxin-like compounds, adding a new dimension to the possible cause of mortalities in A. ocellatum outbreaks. To address this question, extracts prepared from different life stages of the parasite were tested in vitro for cytotoxic effects using two cell lines derived from branchial arches (ABSa15) and the caudal fin (CFSa1) of the gilthead seabream (Sparus aurata), and for hemolytic effects using erythrocytes purified from the blood of gilthead seabream juveniles. Cytotoxicity and a strong hemolytic effect, similar to those observed for Karlodinium toxins, were observed for the less polar extracts of the parasitic stage (trophont). A similar trend was observed for the less polar extracts of the infective stage (dinospores), although cell viability was only affected in the ABSa15 line. These results suggest that A. ocellatum produces tissue-specific toxic compounds that may have a role in the attachment of the dinospores' and trophonts' feeding process.


Assuntos
Dinoflagellida , Doenças dos Peixes , Parasitos , Dourada , Animais , Doenças dos Peixes/epidemiologia , Doenças dos Peixes/parasitologia , Filogenia , Dourada/parasitologia
7.
Chemosphere ; 303(Pt 3): 135198, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-35660050

RESUMO

The presence of microplastics in the aquatic ecosystem represents a major issue for the environment and human health. The capacity of organic pollutants to adsorb onto microplastic particles raises additional concerns, as it creates a new route for toxic compounds to enter the food web. Current knowledge on the impact of pristine and/or contaminated microplastics on aquatic organisms remains insufficient, and we provide here new insights by evaluating their biological effects in zebrafish (Danio rerio). Zebrafish larvae were raised in ZEB316 stand-alone housing systems and chronically exposed throughout their development to polyethylene particles of 20-27 µm, pristine (MP) or spiked with benzo[α]pyrene (MP-BaP), supplemented at 1% w/w in the fish diet. While they had no effect at 30 days post-fertilization (dpf), MP and MP-BaP affected growth parameters at 90 and 360 dpf. Relative fecundity, egg morphology, and yolk area were also impaired in zebrafish fed MP-BaP. Zebrafish exposed to experimental diets exhibited an increased incidence of skeletal deformities at 30 dpf as well as an impaired development of caudal fin/scales, and a decreased bone quality at 90 dpf. An intergenerational bone formation impairment was also observed in the offspring of parents exposed to MP or MP-BaP through a reduction of the opercular bone in 6 dpf larvae. Beside a clear effect on bone development, histological analysis of the gut revealed a reduced number of goblet cells in zebrafish fed MP-BaP diet, a sign of intestinal inflammation. Finally, exposure of larvae to MP-BaP up-regulated the expression of genes associated with the BaP response pathway, while negatively impacting the expression of genes involved in oxidative stress. Altogether, these data suggest that long-term exposure to pristine/contaminated microplastics not only jeopardizes fish growth, reproduction performance, and skeletal health, but also causes intergenerational effects.


Assuntos
Microplásticos , Poluentes Químicos da Água , Animais , Benzo(a)pireno/análise , Ecossistema , Larva , Microplásticos/toxicidade , Plásticos/metabolismo , Polietileno/metabolismo , Poluentes Químicos da Água/análise , Peixe-Zebra/metabolismo
8.
Environ Toxicol Pharmacol ; 90: 103822, 2022 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-35101594

RESUMO

The marine habitat and its biodiversity can be impacted by released pharmaceuticals. The short-term (7 days) effect of 3 commonly used drugs - warfarin, dexamethasone and imidazole - on Senegalese sole (Solea senegalensis) juveniles was investigated. Occurrence of hemorrhages, histopathological alterations, antioxidant status, activity of antioxidant enzymes and expression of genes involved in the xenobiotic response (pxr, abcb1 and cyp1a), were evaluated. The results showed a time and drug-dependent effect. Warfarin exposure induced hemorrhages, hepatocyte vacuolar degeneration, and altered the activity of glutathione peroxidase (GPx) and the expression of all the studied genes. Dexamethasone exposure increased liver glycogen content, altered antioxidant status, GPx and superoxide dismutase activities, as well as abcb1 and cyp1a expression. Imidazole induced hepatocyte vacuolar degeneration and ballooning, and altered the antioxidant status and expression of the tested genes. The present work anticipates a deeper impact of pharmaceuticals on the aquatic environment than previously reported, thus underlining the urgent need for an integrated risk assessment.


Assuntos
Dexametasona/toxicidade , Linguados , Imidazóis/toxicidade , Varfarina/toxicidade , Animais , Antioxidantes/análise , Hemorragia/induzido quimicamente , Fígado/efeitos dos fármacos , Medição de Risco , Transcriptoma , Poluentes Químicos da Água/toxicidade
9.
Ecotoxicol Environ Saf ; 226: 112838, 2021 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-34607190

RESUMO

Persistent and ubiquitous organic pollutants, such as the polycyclic aromatic hydrocarbon benzo[⍺]pyrene (BaP), represent a major threat to aquatic organisms and human health. Beside some well-documented adverse effects on the development and reproduction of aquatic organisms, BaP was recently shown to affect fish bone formation and skeletal development through mechanisms that remain poorly understood. In this work, zebrafish bone-related in vivo assays were used to evaluate the osteotoxic effects of BaP during bone development and regeneration. Acute exposure of zebrafish larvae to BaP from 3 to 6 days post-fertilization (dpf) induced a dose-dependent reduction of the opercular bone size and a depletion of osteocalcin-positive cells, indicating an effect on osteoblast maturation. Chronic exposure of zebrafish larvae to BaP from 3 to 30 dpf affected the development of the axial skeleton and increased the incidence and severity of skeletal deformities. In young adults, BaP affected the mineralization of newly formed fin rays and scales, and impaired fin ray patterning and scale shape, through mechanisms that involve an imbalanced bone remodeling. Gene expression analyses indicated that BaP induced the activation of xenobiotic and metabolic pathways, while negatively impacting extracellular matrix formation and organization. Interestingly, BaP exposure positively regulated inflammation markers in larvae and increased the recruitment of neutrophils. A direct interaction between neutrophils and bone extracellular matrix or bone forming cells was observed in vivo, suggesting a role for neutrophils in the mechanisms underlying BaP osteotoxicity. Our work provides novel data on the cellular and molecular players involved in BaP osteotoxicity and brings new insights into a possible role for neutrophils in inflammatory bone reduction.


Assuntos
Hidrocarbonetos Policíclicos Aromáticos , Peixe-Zebra , Animais , Benzo(a)pireno/toxicidade , Humanos , Larva , Pirenos
10.
Front Nutr ; 8: 719438, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34485367

RESUMO

Osteoporosis is an aging-related disease and a worldwide health issue. Current therapeutics have failed to reduce the prevalence of osteoporosis in the human population, thus the discovery of compounds with bone anabolic properties that could be the basis of next generation drugs is a priority. Marine plants contain a wide range of bioactive compounds and the presence of osteoactive phytochemicals was investigated in two halophytes collected in Brittany (France): the invasive Spartina alterniflora and the native Salicornia fragilis. Two semi-purified fractions, prepared through liquid-liquid extraction, were assessed for phenolic and flavonoid contents, and for the presence of antioxidant, mineralogenic and osteogenic bioactivities. Ethyl acetate fraction (EAF) was rich in phenolic compounds and exhibited the highest antioxidant activity. While S. fragilis EAF only triggered a weak proliferative effect in vitro, S. alterniflora EAF potently induced extracellular matrix mineralization (7-fold at 250 µg/mL). A strong osteogenic effect was also observed in vivo using zebrafish operculum assay (2.5-fold at 10 µg/mL in 9-dpf larvae). Results indicate that polyphenol rich EAF of S. alterniflora has both antioxidant and bone anabolic activities. As an invasive species, this marine plant may represent a sustainable source of molecules for therapeutic applications in bone disorders.

11.
J Cell Biochem ; 122(10): 1556-1566, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34254709

RESUMO

Dual specificity phosphatase 4 (DUSP4), a member of the dual specificity phosphatase family, is responsible for the dephosphorylation and inactivation of ERK, JNK and p38, which are mitogen-activated protein kinases involved in cell proliferation, differentiation and apoptosis, but also in inflammation processes. Given its importance for cellular signalling, DUSP4 is subjected to a tight regulation and there is growing evidence that its expression is dysregulated in several tumours. However, the mechanisms underlying DUSP4 transcriptional regulation remain poorly understood. Here, we analysed the regulation of the human DUSP4 promoters 1 and 2, located upstream of exons 1 and 2, respectively, by the cancer-related transcription factors (TFs) STAT3, FOXA1, CTCF and YY1. The presence of binding sites for these TFs was predicted in both promoters through the in silico analysis of DUSP4, and their functionality was assessed through luciferase activity assays. Regulatory activity of the TFs tested was found to be promoter-specific. While CTCF stimulated the activity of promoter 2 that controls the transcription of variants 2 and X1, STAT3 stimulated the activity of promoter 1 that controls the transcription of variant 1. YY1 positively regulated both promoters, although to different extents. Through site-directed mutagenesis, the functionality of YY1 binding sites present in promoter 2 was confirmed. This study provides novel insights into the transcriptional regulation of DUSP4, contributing to a better comprehension of the mechanisms of its dysregulation observed in several types of cancer.


Assuntos
Fator de Ligação a CCCTC/metabolismo , Fosfatases de Especificidade Dupla/genética , Fator 3-alfa Nuclear de Hepatócito/metabolismo , Fosfatases da Proteína Quinase Ativada por Mitógeno/genética , Fator de Transcrição STAT3/metabolismo , Fator de Transcrição YY1/metabolismo , Apoptose/fisiologia , Sítios de Ligação , Fator de Ligação a CCCTC/genética , Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Fosfatases de Especificidade Dupla/metabolismo , Células HEK293 , Fator 3-alfa Nuclear de Hepatócito/genética , Humanos , Fosfatases da Proteína Quinase Ativada por Mitógeno/metabolismo , Regiões Promotoras Genéticas , Fator de Transcrição STAT3/genética , Fator de Transcrição YY1/genética
12.
Front Cell Dev Biol ; 9: 672424, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34179000

RESUMO

Osteopenia and osteoporosis are bone disorders characterized by reduced bone mineral density (BMD), altered bone microarchitecture and increased bone fragility. Because of global aging, their incidence is rapidly increasing worldwide and novel treatments that would be more efficient at preventing disease progression and at reducing the risk of bone fractures are needed. Preclinical studies are today a major bottleneck to the collection of new data and the discovery of new drugs, since they are commonly based on rodent in vivo systems that are time consuming and expensive, or in vitro systems that do not exactly recapitulate the complexity of low BMD disorders. In this regard, teleost fish, in particular zebrafish and medaka, have recently emerged as suitable alternatives to study bone formation and mineralization and to model human bone disorders. In addition to the many technical advantages that allow faster and larger studies, the availability of several fish models that efficiently mimic human osteopenia and osteoporosis phenotypes has stimulated the interest of the academia and industry toward a better understanding of the mechanisms of pathogenesis but also toward the discovery of new bone anabolic or antiresorptive compounds. This mini review recapitulates the in vivo teleost fish systems available to study low BMD disorders and highlights their applications and the recent advances in the field.

13.
Front Cell Dev Biol ; 9: 642136, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33996798

RESUMO

Keutel syndrome (KS) is a rare autosomal recessive genetic disorder that was first identified in the beginning of the 1970s and nearly 30 years later attributed to loss-of-function mutations in the gene coding for the matrix Gla protein (MGP). Patients with KS are usually diagnosed during childhood (early onset of the disease), and the major traits include abnormal calcification of cartilaginous tissues resulting in or associated with malformations of skeletal tissues (e.g., midface hypoplasia and brachytelephalangism) and cardiovascular defects (e.g., congenital heart defect, peripheral pulmonary artery stenosis, and, in some cases, arterial calcification), and also hearing loss and mild developmental delay. While studies on Mgp -/- mouse, a faithful model of KS, show that pathologic mineral deposition (ectopic calcification) in cartilaginous and vascular tissues is the primary cause underlying many of these abnormalities, the mechanisms explaining how MGP prevents abnormal calcification remain poorly understood. This has negative implication for the development of a cure for KS. Indeed, at present, only symptomatic treatments are available to treat hypertension and respiratory complications occurring in the KS patients. In this review, we summarize the results published in the last 50 years on Keutel syndrome and present the current status of the knowledge on this rare pathology.

14.
Mar Environ Res ; 168: 105309, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33798995

RESUMO

Pharmaceuticals represent a group of emerging contaminants. The short-term effect (3 and 7 days) of warfarin (1 and 10 mg L-1), dexamethasone (0.392 and 3.92 mg L-1) and imidazole (0.013 and 0.13 mg L-1) exposure was evaluated on mussels (Mytilus galloprovincialis). Total antioxidant status, glutathione reductase, glutathione peroxidase (GPx) and superoxide dismutase enzyme activities, and the expression of genes involved in the xenobiotic response (ATP binding cassette subfamily B member 1 (abcb1) and several nuclear receptor family J (nr1j) isoforms), were evaluated. All nr1j isoforms are suggested to be the xenobiotic receptor orthologs of the NR1I family. All drugs increased GPx activity and altered the expression of particular nr1j isoforms. Dexamethasone exposure also decreased abcb1 expression. These findings raised some concerns regarding the release of these pharmaceuticals into the aquatic environment. Thus, further studies might be needed to perform an accurate environmental risk assessment of these 3 poorly studied drugs.


Assuntos
Mytilus , Preparações Farmacêuticas , Poluentes Químicos da Água , Animais , Biomarcadores , Glutationa Peroxidase/genética , Mytilus/genética , Poluentes Químicos da Água/toxicidade
15.
Biomark Med ; 14(8): 639-650, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32613839

RESUMO

Aim: To provide novel data on the expression of DUSP4 transcripts in colorectal cancer (CRC) tissues and to explore their potential as biomarkers. Materials & methods:DUSP4 transcripts expression was determined by quantitative real-time PCR in tissues from 28 CRC patients. Their association with clinicopathological factors and survival analysis was performed. Data from 380 CRC patients available at The Cancer Genome Atlas project were also analyzed. Results: All transcripts were overexpressed in CRC tissues. Variant X1 was the most upregulated and associated with KRAS mutations and poorly differentiated tumor. Overexpression of DUSP4 transcripts could distinguish all tumor stages from normal tissues. Similar results were found in The Cancer Genome Atlas cohort. Conclusion:DUSP4 transcripts have the potential to serve as diagnostic biomarkers for CRC, particularly variant X1.


Assuntos
Biomarcadores Tumorais/genética , Neoplasias Colorretais/genética , Fosfatases de Especificidade Dupla/genética , Regulação Neoplásica da Expressão Gênica , Fosfatases da Proteína Quinase Ativada por Mitógeno/genética , Mutação , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Neoplasias Colorretais/diagnóstico , Diagnóstico Diferencial , Feminino , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Prognóstico , Reação em Cadeia da Polimerase em Tempo Real/métodos
16.
Bone ; 138: 115480, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32534223

RESUMO

The last decade has seen an increased interest in the discovery of compounds with bone anabolic activity to treat skeletal disorders such as osteoporosis and increase the well-being of patients. Due to the many technical advantages over classical rodent systems, zebrafish (Danio rerio) has been increasingly used in screening pipelines, in particular those aiming at identifying osteoactive compounds with pharmacological potential. Because compound osteoactivity is mostly determined in zebrafish through the morphometric analysis of bone structures, image analysis, rather than screening assay implementation, molecule availability and image acquisition, represents a bottleneck to the screening throughput. The absence of auto/semi-automatic tools for image analysis of fish bone structures is also a limitation to a broader usage of zebrafish screening pipelines. We present here ZFBONE (for ZebraFish BONE), an open-source, freely available, user-friendly, rapid and reliable toolset, aiming at accelerating image analysis by automating the morphometric assessment of zebrafish bone structures, but also at increasing data accuracy by reducing operator bias. Tools included in ZFBONE allow users to assess, from 2D images, morphometric parameters of several bone structures (e.g. operculum, caudal fin rays and scales) but also the extent and the intensity of bone-specific colorations. ZFBONE has been developed using the open-source ImageJ software, to make it available to the whole zebrafish research community, but also to have it easily modifiable according to user demands. ZFBONE can also be used toward the standardization of zebrafish screening protocols in academia and industry.


Assuntos
Osteoporose , Peixe-Zebra , Animais , Osso e Ossos/diagnóstico por imagem , Humanos , Processamento de Imagem Assistida por Computador , Software
17.
Int J Mol Sci ; 21(10)2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-32429051

RESUMO

Vitamin K (VK) is a key nutrient for several biological processes (e.g., blood clotting and bone metabolism). To fulfill VK nutritional requirements, VK action as an activator of pregnane X receptor (Pxr) signaling pathway, and as a co-factor of γ-glutamyl carboxylase enzyme, should be considered. In this regard, VK recycling through vitamin K epoxide reductases (Vkors) is essential and should be better understood. Here, the expression patterns of vitamin K epoxide reductase complex subunit 1 (vkorc1) and vkorc1 like 1 (vkorc1l1) were determined during the larval ontogeny of Senegalese sole (Solea senegalensis), and in early juveniles cultured under different physiological conditions. Full-length transcripts for ssvkorc1 and ssvkorc1l1 were determined and peptide sequences were found to be evolutionarily conserved. During larval development, expression of ssvkorc1 showed a slight increase during absence or low feed intake. Expression of ssvkorc1l1 continuously decreased until 24 h post-fertilization, and remained constant afterwards. Both ssvkors were ubiquitously expressed in adult tissues, and highest expression was found in liver for ssvkorc1, and ovary and brain for ssvkorc1l1. Expression of ssvkorc1 and ssvkorc1l1 was differentially regulated under physiological conditions related to fasting and re-feeding, but also under VK dietary supplementation and induced deficiency. The present work provides new and basic molecular clues evidencing how VK metabolism in marine fish is sensitive to nutritional and environmental conditions.


Assuntos
Linguados/crescimento & desenvolvimento , Linguados/metabolismo , Especificidade de Órgãos , Vitamina K Epóxido Redutases/metabolismo , Vitamina K/metabolismo , Sequência de Aminoácidos , Animais , Sequência Conservada , DNA Complementar/genética , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Linguados/genética , Regulação da Expressão Gênica no Desenvolvimento , Regulação Enzimológica da Expressão Gênica , Filogenia , Vitamina K Epóxido Redutases/química , Vitamina K Epóxido Redutases/genética
18.
Mar Biotechnol (NY) ; 22(3): 333-347, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32080776

RESUMO

Teleosts have the ability to regenerate their caudal fin upon amputation. A highly proliferative mass of undifferentiated cells called blastema forms beneath wound epidermis and differentiates to regenerate all missing parts of the fin. To date, the origin and fate of the blastema is not completely understood. However, current hypotheses suggest that the blastema is comprised of lineage-restricted dedifferentiated cells. To investigate the differentiation capacity of regenerating fin-derived cells, primary cultures were initiated from the explants of 2-days post-amputation (dpa) regenerates of juvenile gilthead seabream (Sparus aurata). These cells were subcultured for over 30 passages and were named as BSa2. After 10 passages they were characterized for their ability to differentiate towards different bone cell lineages and mineralize their extracellular matrix, through immunocytochemistry, histology, and RT-PCR. Exogenous DNA was efficiently delivered into these cells by nucleofection. Assessment of lineage-specific markers revealed that BSa2 cells were capable of osteo/chondroblastic differentiation. BSa2 cells were also found to be capable of osteoclastic differentiation, as demonstrated through TRAP-specific staining and pit resorption assay. Here, we describe the development of the first successful cell line viz., BSa2, from S. aurata 2-dpa regenerating caudal fins, which has the ability of multilineage differentiation and is capable of in vitro mineralization. The availability of such in vitro cell systems has the potential to stimulate research on the mechanisms of cell differentiation during fin regeneration and provide new insights into the mechanisms of bone formation.


Assuntos
Nadadeiras de Animais/fisiopatologia , Diferenciação Celular , Regeneração/fisiologia , Dourada , Nadadeiras de Animais/citologia , Nadadeiras de Animais/cirurgia , Animais , Calcificação Fisiológica/fisiologia , Linhagem Celular , Osteoblastos
19.
Zebrafish ; 17(1): 18-26, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31994994

RESUMO

Many anthropogenic chemicals and plastic debris end up in the aquatic ecosystem worldwide, representing a major concern for the environment and human health. Small teleosts, such as zebrafish (Danio rerio) and Japanese medaka (Oryzias latipes), offer significant advantages over classical animal models and are currently used as first-line organisms to assess environmental risks associated with many aquatic toxicants. Toxicological studies require the use of inert materials and controlled conditions. Yet, none of the available commercialized systems is adequate to assess the toxic effect of microplastics, because they contain components made of plastic polymers that may release micrometric plastic particles, leach manufacturing compounds, or adsorb chemicals. The ZEB316 stand-alone housing system presented in this study is meant to be a cost-effective and easy-to-built solution to perform state-of-the-art toxicological studies. It is built with inert and corrosion-resistant materials and provides good housing conditions through efficient recirculation and filtration systems. Assessment of water parameters and fish growth performance showed that the ZEB316 provides housing conditions comparable to those available from commercial housing systems.


Assuntos
Abrigo para Animais , Microplásticos/toxicidade , Oryzias , Testes de Toxicidade/métodos , Poluentes Químicos da Água/toxicidade , Peixe-Zebra , Animais , Monitoramento Ambiental/métodos
20.
Biol Trace Elem Res ; 196(2): 629-638, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31828720

RESUMO

Trace minerals and vitamins are known modulators of bone metabolism, and dietary optimization of these components may improve skeletal development and reduce the occurrence of skeleton deformities in farmed fish. As for larval stages, mineral and water-soluble vitamin nutrition requirements are lacking in research efforts and knowledge is scarce. An in vitro cell system developed from gilthead seabream vertebra and capable of mineralization was used to assess the effect of B vitamins (thiamin and pyridoxine) and trace minerals (copper, manganese, and zinc in a sulfated and chelated form) on cell proliferation and extracellular matrix (ECM) mineralization. Dependent on dose, inhibition of cellular proliferation and/or cytotoxic effects was observed for all nutrients tested and LD50 values were determined: copper, 67.4-69.5 ppm; manganese, 20.9-29.8 ppm; zinc, 37.1-42.8 ppm in sulfated and chelated form respectively; thiamin, 6273 ppm; pyridoxine, 14226 ppm. ECM mineralization was enhanced by mineral (dose and form dependent) and vitamin (dose dependent) supplementation, at non-toxic concentrations below the determined LD50s. This in vitro work confirmed the mineralogenic action of trace minerals and water-soluble vitamins and provided valuable insights for subsequent in vivo nutritional trials.


Assuntos
Minerais/farmacologia , Oligoelementos/farmacologia , Complexo Vitamínico B/farmacologia , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Minerais/química , Dourada , Relação Estrutura-Atividade , Oligoelementos/química , Complexo Vitamínico B/química
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